title: DNA polymerases β and λ do not directly affect Ig variable region somatic hypermutation although their absence reduces the frequency of mutations.
abstract: During somatic hypermutation (SHM) of antibody variable (V) region genes, activation-induced cytidine deaminase (AID) converts dC to dU, and dUs can either be excised by uracil DNA glycosylase (UNG), by mismatch repair, or replicated over. If UNG excises the dU, the abasic site could be cleaved by AP-endonuclease (APE), introducing the single-strand DNA breaks (SSBs) required for generating mutations at A:T bp, which are known to depend upon mismatch repair and DNA Pol η. DNA Pol β or λ could instead repair the lesion correctly. To assess the involvement of Pols β and λ in SHM of antibody genes, we analyzed mutations in the VDJh4 3' flanking region in Peyer's patch germinal center (GC) B cells from polβ(-/-)polλ(-/-), polλ(-/-), and polβ(-/-) mice. We find that deficiency of either or both polymerases results in a modest but significant decrease in V region SHM, with Pol β having a greater effect, but there is no effect on mutation specificity, suggesting they have no direct role in SHM. Instead, the effect on SHM appears to be due to a role for these enzymes in GC B cell proliferation or viability. The results suggest that the BER pathway is not important during V region SHM for generating mutations at A:T bp. Furthermore, this implies that most of the SSBs required for Pol η to enter and create A:T mutations are likely generated during replication instead. These results contrast with the inhibitory effect of Pol β on mutations at the Ig Sμ locus, Sμ DSBs and class switch recombination (CSR) reported previously. We show here that B cells deficient in Pol λ or both Pol β and λ proliferate normally in culture and undergo slightly elevated CSR, as shown previously for Pol β-deficient B cells.
authors.affiliation: Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, MA 01605, United States.
authors.forename: Carol E
authors.initials: CE
authors.lastname: Schrader
authors.forename: Erin K
authors.initials: EK
authors.lastname: Linehan
authors.forename: Anna J
authors.initials: AJ
authors.lastname: Ucher
authors.forename: Barbara
authors.initials: B
authors.lastname: Bertocci
authors.forename: Janet
authors.initials: J
authors.lastname: Stavnezer
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D002478:Cells, Cultured; D053903:DNA Breaks, Double-Stranded; D019951:DNA Polymerase beta / Q000378:metabolism*; D004622:Embryo, Mammalian; D005260:Female; D017353:Gene Deletion; D017578:Immunoglobulin Class Switching / Q000235:genetics; D007074:Immunoglobulin G / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D059645:Mutation Rate; D017354:Point Mutation; D027041:Somatic Hypermutation, Immunoglobulin / Q000235:genetics*
chemical_list: D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; C415367:DNA polymerase beta2; D019951:DNA Polymerase beta
keywords: A:T mutations; AID; Activation induced cytidine deaminase; BER; Base excision repair; CSR; DSB; GC; MMR; Mismatch repair; Pol; SHM; SSB; UNG; activation induced cytidine deaminase; base excision repair; class switch recombination; double-strand DNA break; germinal center; mismatch repair; polymerase; single-strand DNA break; somatic hypermutation; uracil-N-DNA glycosylase